JPS59501745A - Means of removing microorganisms from tissues - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] Means of removing microorganisms from tissues This invention provides a means for removing microorganisms from tissues such as mucous membranes, skin and wound tissue means).

There are unique non-pathogenic and potentially pathogenic microbiota on the skin and mucous membranes. this The presence of proper phase has certain useful functions, and it is said that It can prevent knee formation. Thus, those who carry pathogenic microorganisms are latent pathogen carriers because pathogenic microorganisms from individuals can be transferred to tissue wounds. It is.

Microbial terms include Staphylococcus, Streptococcus and Pneumococci. Gram-positive bacteria such as various species of mycobacteria, e.g. Gram-positive rods such as M. lepres, various species of Enterobacteriaceae, e.g. Escherichia coli, talebcilla, proteune, enterobacter or Gram-negative bacteria such as Vibrionase or Pseudomonase as well as Gonococci This includes gram-negative cocci such as. This word also refers to batateroides, Anaerobic bacteria such as Peptococci and Peptostrep Also includes anaerobic cocci such as Tococcii, as well as Nupirile and Nupirochetes. shall be held. Additionally, fungi such as Candida, such as Candida albicans, , and Dermot 7ites, and diseases such as Endamoeba histolytica. amoeba, and also includes Giardia lampria and Leishmania shall be taken as a thing.

Methods for removing microorganisms from tissues have previously differed depending on the tissue and type of microorganism. Ru. To eliminate pathogenic microorganisms from mucous membranes and skin tissue, use disinfectants or antibiotics. It is often used. The disadvantage of this method is that the eigenphase and its functions are also removed. Ru. Another disadvantage is that disinfectants are cytotoxic, and today many pathogenic strains are treated with a large number of antibiotics. This means that they have acquired resistance to the substance. Elimination of pathogenic microorganisms from wound tissue The removal can be done in several ways. One method is to use a disinfectant or antibiotic treatment. direct attack of microorganisms by microorganisms, but this is not effective and has the above-mentioned disadvantages in the indigenous phase. Tarasu. Another method is mechanical cleaning of the wound tissue with a moist saline dressing. lack of this In particular, tissue healing is inhibited and such dressings must be changed very frequently. This is something that needs to be done.

The latter is an example of a more indirect method of removing pathogenic microorganisms from wound tissue. . Secretions from wound tissue are excellent substrates for microorganisms, and their reduction in Therefore, the growth of microorganisms is also reduced. During this time, another method using the tangential method is 1 e.g. Wounds made of a suitable polymer, which absorbs secretions from the wound tissue. This is the process. Some of these secretion-absorbing polymers appear to have complex bonds. (so-called iodophor), which simultaneously absorbs secretions and disinfects them. will be accomplished.

This invention relates to a means of removing microorganisms from tissues without significantly affecting the indigenous phase. It is something to do.

Staphylococcus aureus is a major cause of primary wound infections and postoperative infections. certain types of streptococci, such as the so-called AlC and group 0 beta-hemolytic streptococci. It is a gram-positive coccus. Pathogenic Staphylococci can be found anywhere in the environment and It is present everywhere in the hospital and 80% of the staff can be carriers of the pathogen. yellow Staphylococcus aureus is a staphylococcus aureus that is found in certain areas of the body, especially in the axilla. Skin area, pelvis 4 Builds a colony at the bottom and inside the nose. Even dry eczema is often white and Staphylococcus aureus is present. Streptococcus, the bacterium that causes tonsillitis Cus pyogenes (group A Streptococcus) is very often found in nurses and present in the patient. Nosocomial infections can be caused by external sources (e.g. from staff to patients) or are endogenous (infected skin or mucous membranes by patients who have these microorganisms or their own wounds). ), and this is becoming a major problem. To reduce the frequency of this infection , Note 4: Classical prevention such as careful preoperative cleaning and bandaging infected eczema in a separate room. Measures have been taken.

The first step in most infections is the attachment of microorganisms to tissue. microorganisms blood clots or concretions exposed directly to surface components of epithelial cells or during epithelial wounds. It specifically binds to extracellular molecules in synaptic tissue.

This invention is based on a completely new principle for removing microorganisms from tissues. , its principle is based on the inhibition of microbial binding to mucosal, skin and wound tissue proteins. It is the foundation.

Pathogenic Staphylococci have a variety of immune systems on their cell surface that are found in many animal species. A protein that specifically binds to the complement binding site (Fo site) of epiglobulin. A). This binding specificity is due to immunotenuating and solid phase-bound proteins. It is used to remove antibodies from the extracorporeal circulation by In-A.

It is also known that Staphylococcus agglutinates with normal serum from various animal species. It is. This reaction involves the specific binding of fibrin/fibrinogen to the bacterial surface. This is due to the

Receptors similar to those present on the surface of Staphylococci or other microorganisms It was discovered that it exists. Examples are isolated from wound infections and other types of infections. The results of repeated research on Staphylococcus and Streptococcus strains , for example, the ability to bind fibronectin has been found to contribute to microbial virulence. Ta. For example, Staphylococcus aureus and coagulase negative staphylo In coccal infections, this binding ability may allow initial tissue colonization. contributes more to toxicity.

Surprisingly, the protein or its 7 fragments may be optionally bound to a carrier. or any suitable means of removing microorganisms from the residue or tissue. It was found that That is, the present invention can be applied to In the removal of microorganisms from tissues, proteins that can bind to microorganisms are still at least one fragment or residual portion, and optionally the protein or or the means comprising the carrier to which the fragment or residual part is attached. It is something that

Suitable proteins for use in this invention include fibrinogen, fibronectin, Immunoglobulin, albumin, haptoglobin, beta-2-microglobulin , collagen, laminin, entactin, Matahachondroi cutin. Sara Additionally, the seven fragments of these proteins, as well as the remaining classes, are also suitable. These 7 The lagnont or the remainder consists of molecules 1sx1o2 to 105, preferably 5x10 There may be 2 to 5 x 104 peptides.

Suitable carriers for use in this invention include acarose or its derivatives, starch or its derivatives, cellulose or its derivatives, dextran or its derivatives, or or a polymeric material such as alginate. Additionally, bandages or tampons Also suitable are fibrous materials forming part of such absorbent products.

Proteins or fragments thereof (head) or other portion(s) thereof are known per se. For example, divinyl sulfone, epichlorohydrin, carboxylic acid or by transnumination or cyanogen bromide crosslinking. or binding of fibronectin to gelatin, or glycoprotein to lectin. biospecific interactions known per se, such as antigen binding or antibody binding; carbohydrate moieties, primary amine groups, sulfhydryls or carbons in proteins. It can be attached to a carrier through a covalent bond via a carboxy group.

Proteins or their 7 fragments or residual parts can be directly absorbed into a carrier. You can also do that.

Further, this invention will be explained by the following examples.

Example 1 The human lumbar skin area was washed with saline. Next, Staphylococcus a 107 and 109 were applied to several skin areas. centre Sephalone (trademark) 4B gel bound to fibronectin was added to a thickness of 2-3 mm. Newman stock suitable as a layer of 8 applied to the area of use. Fibrolectin is activated by cyanogen bromide. Incubate 10 Mg of fibronectin for 2 hours at room temperature with gentle agitation. Binds to Sepharose™ 4B gel activated with cyanogen bromide by I let it happen. After terminating the reaction by adding 1M glycine solution to the reaction mixture, the incubator Bation continued for an additional 30 minutes. The gel was thoroughly washed before use. control gel and Then apply pure Sepharose™ 4B to another Newman strain applied skin area in the same way. used. The entire skin area was dried with a fan (cold air). After 20-30 minutes, test gel and control gel were removed with a sterile scalpel. Then apply new gels and repeat the operation. The treatment was repeated for a total of three times over a period of approximately 2 hours.

After the completion of the procedure, the Newman strain was almost completely eliminated by culturing (only one spot Bacterial colonies were observed in the skin cells treated with the control gel. Cultivation of the area showed only a slight decrease in the number of bacteria.

Example 2 Staphylococcus aureus (strain v'8) was used as a substitute for the Newman strain. Example 1 was repeated. A control gel of a similar type was used. For culture after treatment is completed, Growth of 8 strains of Shimokana was observed compared to the Newman strain of Example 1.

Example 3 Example 1 was repeated using fibrinogen in place of fibronectin. Sef Binding of fibrinogen to Wallose™ 4B gel was performed using the fibrinogen of Example 1. The same procedure as for bronectin was performed. Pure Sepharose(TM) as control gel 4B gel was used.

By culturing after completing the operation, ■8 strains were almost completely removed (only one colony However, in the culture of the skin area to which the control gel was applied, A slight decrease in the number of bacteria was observed.

These examples show that the Newman strain has fibronectin receptors and The v8 strain binds extremely weakly to fibronectin, but does not bind fibrinogen. It turned out that he had a scepter. Furthermore, strains with Fn receptors have specific It is clear that this will be excluded.

Example 4-19 Staphylococcus aureus 5A113 strain 0 Cowan 1 strain and Group A Nutreptococcus H15757 Example 1 was repeated using strains 1 and 11270.

The proteins used are albumin, immunoglobulin G1 fibrinogen, and and fibronectin, and like the fibronectin in Example 1, these are It was bound to a Fallof 4B gel.

The procedure of Example 1 was carried out, and then cultured to produce various microorganisms and proteins. We obtained the results described below.

Using fibronectin, Staphylococcus aureus (SA113 The culture of Al. Using Bumin, Immunoglobulin G and Fibrinogen respectively, this A positive culture result was obtained for the strain.

Using immunoglobulin G, fibrinogen and fibronectin, respectively Cultures of Staphylococcus aureus (Cowan strain 1) were negative; , positive culture results were obtained for this strain using albumin.

Using albumin and fibronectin, respectively, group A streptococci Cultures of strains (H15757) were negative, while immunoglobulin C and fibrin Positive culture results were obtained for this strain using linogen, respectively.

Culture of group A nutreptococcus (strain 11270) using fibrinogen was negative, while albumin, immunoglobulin G and fibronectin were used. positive culture results were obtained for this strain.

In all cases, cultures of the skin area to which the control gel was applied gave positive culture results. or obtained.

Example 20 Hospitals certified as colonized by antibiotic-resistant Staphylococcus aureus The staff were divided into two groups, with equal numbers in each group (test group and control group, respectively). 2 sets of all Samples from patients repeatedly showed positive culture results. The test group was Treat with a cotton swab soaked in solution, insert the swab into the nostril and hold for 2 hours, then remove the fibrous material. It was replaced with a new cotton swab soaked in nectin solution. This operation is processed a total of 3 times. Naruma Tekushi replied. The control group was treated similarly using a non-soaked cotton swab.

Samples obtained from persons in the control group after completion of the procedure showed positive cultures, whereas samples obtained from persons in the test group The sample obtained from the laboratory was completely negative.

Example 21 The body hair of a mouse (20-301, CBA strain) was shaved and a 1 x 10 area was placed according to international standards. 2nd or 3rd degree burns (ethanol flame, 2 minutes). In this example, Taphylococcus aureus (Cowan 1 strain) was used as a microorganism. salt Prepare a suspension solution containing 10 microorganisms per ml of water (1 ti of phosphoric acid) and A 0.5 m/ml solution was applied to the burn area. The mice were then divided into test and control groups. Divided into. Mice in the test group received fibrous gel bound to Sepharose™ 4B gel. Mice in the control group were treated with nectin, while mice in the control group were treated with pure Sepharose™ 4B. did. The treatment is applied to the burn area as a gel or an amalgamated layer approximately 2-3 fi thick. I did it like this. After removing one layer by scrubbing with a scalpel, apply a new layer and process. was repeated until a total of three layers were applied and removed. Then sample from the burn area were taken and cultured.

Samples from test group mice showed negative cultures (i.e. 1-3 colonies) after 1 day.

Samples from control group mice were cultured positive even after 3 days (3 or more colonies until fused growth) showed that.

Example 22 The same type of mouse used in Example 21 was used.

Staphylococcus capitis L instead of Staphylococcus aureus The procedure of Example 21 was repeated except that K499 was used.

Samples from test group mice had positive cultures on day 2, while samples from control group mice had positive cultures at day 2. The culture was still positive on day 1.

Example 23 The same type of mouse used in Example 21 was used.

Staphylococcus nemoritake instead of Staphylococcus aureus The procedure of Example 21 was repeated except that the same solution was used.

Samples from both test and control group mice showed positive cultures on day 3.

Example 24 The same procedures and strains as in Example 2-1 were used, and the mice were of the same type, but the test group mice were treated with fibrinogen bound to Sepharose™ 4B gel. Ta.

Samples from test group mice had positive cultures on day 2, while samples from control group mice had positive cultures at day 2. The culture was positive on day 1.

Example 25 Using the same procedure and strain as in Example 22, the mice were also of the same type and rarely in the test group. Mice were treated with fibrinogen bound to Sepharose™ 4B gel. .

Samples from both test and control group mice showed positive cultures after 4 days.

Example 26 Using the same procedures and strains as in Example 23, the mice were also of the same type or different from the test group. Mice were treated with fibrinogen bound to Sepharose™ 4B gel. Ta.

Samples from both test and control group mice showed positive cultures after 4 days.

From the above examples it is clear that different microorganisms bind to different proteins.

Each bond between the microorganism and the protein was confirmed by in vitro experiments. This is for each Microorganisms can be incubated with each protein and the latter labeled with radioactive iodine. It was carried out by After the incubation is complete, the radioactive label bound to each microorganism is The amount of protein was measured. When microorganisms are removed by protein treatment (culture negative sample) In vitro experiments have shown that each microorganism binds to a radioactively labeled protein. , on the other hand, if microorganisms were not removed by protein treatment (culture-positive samples), in vitro In the experiment, no binding between the corresponding microorganism and the radiolabeled protein was observed.

Proteins can also be used in dressings or in tissues to remove microorganisms from wounds or other tissues. It can be bonded to a fibrous material that forms part of an absorbent product such as a pop. protein Dressings that combine quality are applicable to wound treatment.

Protein-bound tampons are used, for example, for absorption through the surgical opening during surgery. It can also be used as a menstrual tampon, and is the cause of so-called tampon disease. It can be used to remove microorganisms.

The means of the invention also provides for the wound cavity created at the opening of a pyoma, such as an anal pyoma. It can be used to remove microorganisms inside, in which case it can be used to fill the cavity and block the abscess. In order to prevent stomach. Such materials may include, for example, fibers contained in hydrophobic materials or in the form of granules. It may be an absorbent material. In addition, the protein is used to remove microorganisms in local infections. It may be liquid engraving.

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Claims (1)

[Claims] 1. In a means for removing microorganisms from tissues such as mucous membranes, skin and wound tissue, Proteins or fragments thereof that can bind to microorganisms (C) or residual parts (C) ) and optionally the above protein(s) or a fragment thereof. (class) or the remaining portion □□□) is bonded to a carrier. means. 2. If the protein or its fragment(s) or residual portion(s) is Linogen, fibronectin, immunoglobulin, albumin, haptoglobin , beta-2-microglobulin, collagen, laminin, entuftin or The means according to claim 1, characterized in that it is chondronectin. 3. 7 fragments (class) or residual parts (class) of proteins (class) or fibrils Nogen, fibronectin, immunocloprin, albumin, haptoglobin, beta-2-microcloprine, collagen, laminin, entuftin or Claim 1 characterized in that it is derived from andronectin. Means described in section. 4 Fragment(s) or residual portion(s) or molecules 1t5X102-105 The means according to claim 3, which is a peptide. 5. The carrier is agarone or its derivative, tenbun or its derivative, cellulose dequinutran or its derivatives, still alginate Any one of claims 1 to 4, characterized in that it is a polymer material such as The means described in. 6. A fibrous material that is either a carrier or part of an absorbent product such as a dressing or a tampon. As described in any one of claims 1 to 4, characterized in that means of. 7. Protein(s) or its 7 fragment(s) or residual portion(s) are Vinyl sulfone, epichlorohydrin, and carbodiimide produce transamines. tion, via cyanogen bromide cross-linking, or by biospecific interactions. Any one of claims 1 to 6, characterized in that it is linked/bonded to the body. Means of description. 8. Protein(s), its fragment(s) or residual portion(s) or carrier Claims 1 to 6, characterized in that: 9 Proteins such as Chin or Fundronectin or their 7 fragments or in the residual class of microbial removal from tissues such as mucous membranes, skin and wound tissue. use. or the remainder portion(s) is bound to or absorbed by a carrier. The proteins or fragments thereof according to claim 9, or Use of residual portion(s). I product 11g59-501745 (2)